Staphylococcus aureus is a major cause of invasive human infections, including bacteremia, endocarditis, pneumonia, and wound infections. Methicillin-resistant S. aureus (MRSA) are endemic in hospitals, and community-associated MRSA strains are spreading worldwide, posing a major global challenge [1-3]. There is an urgent need for a vaccine to prevent staphylococcal disease. Several vaccines have been tested in clinical trials, but capsular polysaccharide (CP) conjugates, individual protein antigens, and monoclonal antibodies (mAbs) to lipoteichoic acid have failed at various developmental stages, underscoring the need for novel vaccines with broader efficacy [4-6]. S. aureus vaccines that elicit both humoral and cell mediated immune responses are currently under evaluation [7], and both alpha toxin (Hla) and CPs are key antigens under consideration for inclusion in a multi73 component vaccine.
Serotype 5 (CP5) or serotype 8 (CP8) capsules are produced by ˜75% of S. aureus clinical isolates, and CP antigens are critical for survival in the blood of infected animals [8, 9]. Capsular antibodies are opsonic, mediating uptake and killing of staphylococci by human neutrophils [8]. Hla is a secreted pore forming toxin to which lymphocytes, macrophages, alveolar epithelial cells, pulmonary endothelium, and erythrocytes are sensitive [10]. A genetically detoxified protein (HlaH35L) is defective in pore formation, and antibodies to HlaH35L neutralize the lytic activity of native Hla [11]. Immunization with HlaH35L protects mice against lethal staphylococcal pneumonia, lethal peritonitis, and skin infections [12-14].
Immunization with conserved staphylococcal protein antigens glycosylated with CPs may be an elegant and efficient strategy to prevent S. aureus infections, limiting the numbers of individual vaccine components that need to be prepared and individually purified. Such an approach is feasible through the development of a novel Escherichia coli N-linked glycosylation technology [15, 16], wherein O antigens are transferred to specific sites within a protein carrier by the oligosaccharyltranferase PglB [15-17]. In contrast to chemically conjugated vaccines, bioconjugate vaccines are homogenous with a defined molecular structure, and the protein and glycan components are kept in native conformations, avoiding denaturation of essential B-cell epitopes [18]. The product contains peptide and covalently linked sugar epitopes from the same organism, thereby broadening its efficacy against numerous manifestations of microbial disease. We have prepared glycoconjugate vaccines comprised of CP5-Pseudomonas aeruginosa exoprotein A (Epa), CP8-Epa, and CP5-Hla and evaluated their protective efficacy against bacteremia and lethal pneumonia in mice. Whereas CP5-Epa and CP8-Epa significantly reduced bacteremia, the CP5-Hla bioconjugate vaccine protected against both bacteremia and lethal pneumonia.